Binding of DNA-bending non-histone proteins destabilizes regular 30-nm chromatin structure

Abstract

Why most of the in vivo experiments do not find the 30-nm chromatin fiber, well studied in vitro, is a puzzle. Two basic physical inputs that are crucial for understanding the structure of the 30-nm fiber are the stiffness of the linker DNA and the relative orientations of the DNA entering/exiting nucleosomes. Based on these inputs we simulate chromatin structure and show that the presence of non-histone proteins, which bind and locally bend linker DNA, destroys any regular higher order structures (e.g., zig-zag). Accounting for the bending geometry of proteins like nhp6 and HMG-B, our theory predicts phase-diagram for the chromatin structure as a function of DNA-bending non-histone protein density and mean linker DNA length. For a wide range of linker lengths, we show that as we vary one parameter, that is, the fraction of bent linker region due to non-histone proteins, the steady-state structure will show a transition from zig-zag to an irregular structure-a structure that is reminiscent of what is observed in experiments recently. Our theory can explain the recent in vivo observation of irregular chromatin having co-existence of finite fraction of the next-neighbor (i + 2) and neighbor (i + 1) nucleosome interactions.

Fig 1

(a) When the DNA (yellow chain) is wrapped around the histone octamer (blue), we can represent the direction of the DNA segments that enters and exits the histone core as two vectors as shown here. The relative orientations of these entry/exit vectors will influence the local structure of the chromatin. (b) Linker DNA is typically rigid and straight. Rigid linker DNA and restricted orientations of entry/exit DNA segments will promote a zig-zag structure. (c–d): Schematic diagram describing the models: (c) Bead-spring model—DNA is modelled as a polymer made of beads of type-1 (yellow). 14 DNA-beads wrap around the histone-octamer bead (type-2, blue) to form a nucleosome. The picture also depicts linker histone H1 constraining entry/exit DNA beads (black spring), non-histone protein bending the linker region (red spring), and rigid linker regions when it is free of non-histone proteins. (d) Equivalently, nucleosome is considered as a bead that constraints the entry/exit DNA segments at an angle α_n. (e) FRC model—chromatin is modelled as a long 3D chain of N vectors. Yellow vectors represent linker DNA segments, which make a small angle θ_d with respect to its neighbor vector (in the absence of a non-histone protein); blue arrows represent histone-bound DNA having an angle θ_n = π − α_n between them. When a non-histone protein is present in the linker region, yellow tangents make a relative angle of θ_p as shown.

Fig 2

(a–b): Snapshots of chromatin from BD simulations showing DNA (yellow) and nucleosomes (blue and green) [55]. (a) In the absence of non-histone proteins; a zig-zag structure is seen. (b) In the presence of non-histone proteins that bend linker DNA (density ρ = 0.5). DNA-bending brings neighbouring nucleosomes closer to each other mixing the blue and green. This destroys the zig-zag nature where, typically, next neighbors (same-color nucleosomes) are closer than the neighbors (different colors). Both have inter-nucleosome interaction with ${\tilde{k}}_h=50$. (c–d): I(k) from BD simulations. (c) Without (${\tilde{k}}_h=0$) and with inter-nucleosome interactions (${\tilde{k}}_h=10,30,50$) and in the absence of any non-histone protein. Here I(k) peaks at k = 2 indicating the formation of zig-zag structure for each ${\tilde{k}}_h$. (d) Without (${\tilde{k}}_h=0$) and with inter-nucleosome interactions (${\tilde{k}}_h=10,30,50$) in the presence of non-histone proteins(ρ = 0.5). Here the peaks at I(1) imply that neighboring nucleosomes are geometrically close to each other, and the zig-zag structure is dismantled in the presence of non-histone proteins.

Fig 3. BD simulation with two specific DNA-bending geometries corresponding nhp6 and HMG-B.

(a) I(k) for proteins having a size of 20bp (one angle involving 2 bonds in the model) with bending angle = 120° (blue) and 90° (green). The red curve is the control when there is no DNA-bending protein. (b) For proteins having a size of 30bp (one angle involving 3 bonds in the model; see S1 Text). The plots are for ${\tilde{k}}_h=50$ and ρ = 0.5 and with nucleosomes as angles α_n = 60° done using LAMMPS [50]. In all the cases, the presence of DNA-bending proteins shifts the peak away from k = 2 indicating the destruction of zig-zag.

Fig 4. Snapshots of chromatin simulated using the FRC model in 2D showing linker DNA (red) and nucleosomes (blue).

The linker length is ≈ 42 bp. (a)In the absence of any non-histone protein, we get a nice zig-zag-like structure. (b)In the presence of non-histone proteins where each linker region has a probability of 0.24 for non-histone protein to bind. The presence of non-histone proteins is modeled as a bend in the linker region. The non-histone proteins (not marked separately) are visible as sharp angles between two neighboring blue dots.

Fig 5. From 3D FRC model with 2000 nucleosomes.

(a) I(k) without any non-histone proteins (blue curve) and with different densities of non-histone proteins: ρ = 0.1 (yellow), ρ = 0.3 (pink), and ρ = 0.5 (green). The peak at k = 2 is shifted to k = 1 on increasing ρ. Here, the bending angle of the non-histone protein is θ_p = 90° representing nhp6 and HMG-B. Inset: the peak location (k value at which I(k) is peaked), k_peak, for different ρ. This indicates a transition from a zig-zag to an irregular structure. (b) The same as (a) but with θ_p chosen randomly from 90°–135° (c) A phase diagram obtained by varying the linker length and the density of proteins in the linker region (θ_p as in (b)). The red dots represent the values of the parameters at which the transition from zig-zag to irregular structure happens (i.e., parameter values at which I(1) = I(2)). (d) A similar phase diagram obtained by varying the bending angle (representing different proteins) and the protein density (linker length = 42bp).